Impact of Several Factors on the Diagnostic Interpretability of Coronary Computed Tomographic Angiography Using a 256-Detector Row CT Scanner

Purpose: To explore the impact of patient-related, vessel-related, image quality-related and cardiovascular risk factors on CCTA interpretability using 256-detector row Computed Tomography (CT). Methods: One hundred ten patients who underwent CCTA and Invasive Coronary Angiography (ICA) were consecutively retrospectively enrolled from January 2018 to October 2018. Using ICA as the reference standard, ≥50% diameter stenosis was defined as the cut-off criterion to detect the diagnostic performance of CCTA. Diagnostic reproducibility was investigated by calculating the interrater reproducibility of CCTA. Multiple logistic regression models were performed to evaluate the impact of 14 objective factors. Results: A total of 1019 segments were evaluated. The per-segment sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of CCTA were 76.8%, 93.7%, 91.2%, 67.8% and 95.9%, respectively. The per-segment diagnostic reproducibility was 0.44 for CCTA. Regarding accuracy, a negative association was found for stenosis severity, calcium load and hyperlipidaemia. Regarding sensitivity, calcium load and diabetes mellitus (DM) were positively related. Regarding specificity, a negative correlation was observed between stenosis severity and calcium load. Regarding interrater reproducibility, stenosis severity and calcium load were negatively associated, whereas male sex and the signal-to-noise ratio (SNR) were positively related (all p<0.05). Conclusion: Per-segment 256-detector row CCTA performance was optimal in stenosis-free or occluded segments. Heavier calcium load was associated with poorer CCTA interpretability. On the one hand, our findings confirmed the rule-out value of CCTA; on the other hand, they suggested improvements in calcium subtractions and deep learning-based tools to improve CCTA diagnostic interpretability.

Reduced Kidney Function and Relative Hypocalciuria—Observational, Cross-Sectional, Population-Based Data

This observational, cross-sectional, epidemiological analysis investigated relationships of kidney function to urine calcium and other variables. The analyses targeted two population-based samples of adults (Gubbio study and Moli-sani study: n = 3508 and 955, respectively). Kidney function was assessed as estimated glomerular filtration rate (eGFR). Calcium/creatinine ratio (Ca/Cr) was used as index of urinary calcium in timed overnight urine under fed condition (Gubbio study), morning urine after overnight fast (Gubbio study), and first-void morning urine (Moli-sani study). Moli-sani study included also data for glomerular filtered calcium load, tubular calcium handling, and serum phosphorus, parathyroid hormone, 1,25-dihydroxyvitamin D, calcium, and 25-hydroxyvitamin D. eGFR positively and independently related to Ca/Cr (p < 0.001). In multivariate analyses, eGFR lower by 10 mL/min × 1.73 m2 related to overnight urine Ca/Cr lower by 14.0 mg/g in men and 17.8 mg/g in women, to morning urine Ca/Cr lower by 9.3 mg/g in men and 11.2 mg/g in women, and to first-void urine Ca/Cr lower by 7.7 mg/g in men and 9.6 mg/g in women (p < 0.001). eGFR independently related to glomerular filtered calcium load (p < 0.001) and did not relate to tubular calcium handling (p ≥ 0.35). In reduced eGFR only (<90 mL/min × 1.73 m2), low urine Ca/Cr independently related to low serum 1,25-dihydroxyvitamin D (p = 0.002) and did not relate to hyperphosphatemia, high serum parathyroid hormone, or hypocalcemia (p ≥ 0.14). Population-based data indicated consistent associations of lower kidney function with lower urine calcium due to reduction in glomerular filtered calcium. In reduced kidney function, relative hypocalciuria associated with higher prevalence of low serum 1,25-dihydroxyvitamin D.

EFFECT OF CALCIUM LOAD ON HEART FUNCTION, MPTP OPENING IN SITU AND UCP2/3 MRNA EXPRESSION IN THE HEART OF TRAINED RATS

We have studied the effect of calcium load (1.7 to 15 mmol/l in perfusate) on isolated heart function, mitochondrial factor release (as a marker of mitochondrial permeability transition pore, MPTP), and cardiac uncoupling proteins (UCP2/3) mRNA expression in untrained and trained rats (swimming for 4 weeks). It was found that the improvement in the isolated heart function of trained rats was accompanied by an increase in the expression of UCP3, but not UCP2. A gradual increase of the calcium content in the perfusate led to an increase in contractile function, more pronounced in trained rats. However, 10 mmol/l and higher concentration of calcium led to arrhythmia and drastic decrease in contractility of isolated heart more obvious in untrained rats. Swimming course prevented the calcium-induced release of mitochondrial factor exerting a stabilizing effect on mitochondrial membranes which was, however, diminished by a nitric oxide synthesis blocker (L-NAME). We have found that UCPs genes expression is calcium-sensitive: an increase in UCP3 mRNA at 5 mmol of calcium and a sharp decrease in UCP2/3 expression at 12.5 mmol/l of calcium in perfusate in both trained and untrained rats indicating the participation of UCPs in the regulation of calcium homeostasis. Our data suggest that the calcium load may serve as a test for in situ MPTP titration. Activation of UCPs together with up-regulated nitric oxide may play a protective role against increasing extracellular calcium inhibiting MPTP formation during physical trainings.

Valvular calcium load assessment for predicting postprocedural paravalvular leakage: a comparison between surgical aortic valve replacement versus transcatheter implantation

Background Calcification of native aortic valve is a well known variable causing paravalvular leakage (PVL) following transcatheter aortic valve implantation (TAVI). However, there is currently no evidence to support the fact that patients with high calcium load could be better treated with surgical aortic valve replacement (SAVR). Purpose To assess the utility of preoperative assessment of valvular calcium load through computed tomography in patients affected by severe aortic valve stenosis undergoing SAVR or TAVI. Methods Between June 2016 and June 2018, 109 candidates for isolated SAVR through minimal invasive access underwent preoperative contrast enhanced multidetector computed tomography (MDCT) for the assessment of valve and aortic calcifications. Calcium load was quantitatively measured using a dedicated software in three regions on interest (aortic valve [AV], left ventricular outflow tract [LVOT] and device landing zone [DLZ], which is the sum of the earlier 2). Clinical, echocardiographic, and MDCT variables were collected and compared to a sample population of 107 patients that underwent TAVI (87 transfemoral, 20 transapical) for native aortic valve stenosis in the same period of time, in the same institution. A univariate and multivariate logistic regression analysis were performed on the whole study population to assess risk factors for the onset of postoperative PVL (any grade, defined as ≥ mild) at discharge. Results The two study groups were significantly different in terms of age (71.9±5 in SAVR; 81.5±6 in TAVI), gender (36% female in SAVR vs 51% in TAVI), Euroscore II (1.9%±0.8 in SAVR; 4.8%±2.7 in TAVI), annulus perimeter (79.5mm±8.2 in SAVR; 61.8mm±30.5 in TAVI), baseline ejection fraction (57%±8 in SAVR; 51%±12 in TAVI) and severe pulmonary hypertension (2%±13 in SAVR; 26%±44 in TAVI). Calcium load was not different between groups (DLZ 1066 mm3±716 vs 955mm3±639; total calcium in AV 987 mm3±678 vs 879 mm3±601; total calcium in LVOT 78 mm3±130 vs 77 mm3±100). 30-days-mortality was 1.8% in SAVR and 5.6% in TAVI group (p=0.17). At discharge, incidence of all grades PVL was 5.5% in SAVR (0.9% trace, 3.6% mild, 0.9% moderate) and 41% in TAVI group (12.1% trace, 25.2% mild, 3.7% moderate) (p&lt;0.05). On logistic regression on the whole study population, DLZ calcium (OR 1.1, 95% CI 1–1.2 for 100 mm3, p=0.003) and the use of TAVI (OR 24, 95% CI 7.7–78, p&lt;0.001) were identified as independently associated with the onset of PVL. Conclusions Aortic valve calcifications are a risk factor for the onset of PVL for both TAVI and SAVR. Nevertheless, the risk increases considerably with the use of TAVI. A deeper anatomical analysis of preoperative MDCT could improve the treatment selection and the outcome of patients affected by aortic valve stenosis. Funding Acknowledgement Type of funding source: None

Genetic variants in calcium regulatory cardiac genes and their contribution to Takotsubo syndrome

Background and purpose Takotsubo syndrome (TTS) is characterized by an acute left ventricular dysfunction similar to a myocardial infarction (MI) in the absence of coronary artery stenosis. Patients show symptoms similar to the acute MI with increased biomarkers and blood serum catecholamines. Recently, we developed a patient-specific TTS stem cell model and identified a higher sensitivity to catecholamine-induced stress. Furthermore, familial TTS cases and genetic studies point to a genetic predisposition. The purpose of this study was to analyze a genetic predisposition by characterizing genetic variants in genes associated with cardiac pathologies and their impact on calcium homoeostasis in TTS. Methods and results Whole exome sequencing analysis of a TTS patient discovered 2 missense AHNAK variants in its C-terminal domain and in addition the missense variant F189L in the calcium buffering calsequestrin 2 gene (CASQ2). AHNAK is a 700kDa big nucleoprotein and is involved in the β-adrenergic regulation of the cardiac calcium channel Cav1.2. 3-month old TTS-iPSC-derived cardiomyocytes (CM) were generated and the variants were confirmed by sequencing. We found AHNAK higher expressed in TTS-iPSC-CMs compared to control, whereas no expression alteration was observed for Cav1.2. Since AHNAK is described to act as a repressor towards Cav1.2, which is relieved under β-adrenergic stimulation, we analyzed the effect of AHNAK variants on a potential co-localization and interaction between both proteins. AHNAK and Cav1.2 were shown to co-localize in the cytoplasm as well as the membranes and co-immunoprecipitation experiments confirmed an interaction of AHNAK and Cav1.2 in all tested control- and TTS-iPSC-CMs. On a functional level, we were able to show by patch clamp analysis that Cav1.2 calcium currents are significantly increased in TTS-iPSC-CMs compared to control. The influence of CASQ2-F189L on sarcomeric reticulum (SR) calcium load was analyzed by epifluorescence microscopy using FURA4 and caffeine-applications. We found significantly decreased SR calcium content with an increased fractional release during systole in TTS-iPSC-CMs. To test, whether these variants are the main reason for altered interaction of AHNAK and Cav1.2, calcium currents or SR calcium load in TTS need to be proven in the future by using CRISPR/Cas9-rescued AHNAK/CASQ2 lines. Conclusion Here we show the cardiac functional consequences of AHNAK and CASQ2 missense mutations in TTS-iPSC-CMs with regard to calcium currents and SR calcium load. These results show that AHNAK and CASQ2 variants may predispose to TTS and enable a new therapeutic option for TTS. Funding Acknowledgement Type of funding source: Foundation. Main funding source(s): Else Kröner-Fresenius Foundation

Coupling of NMDA receptors and TRPM4 guides discovery of unconventional neuroprotectants

Excitotoxicity induced by NMDA receptors (NMDARs) is thought to be intimately linked to high intracellular calcium load. Unexpectedly, NMDAR-mediated toxicity can be eliminated without affecting NMDAR-induced calcium signals. Instead, excitotoxicity requires physical coupling of NMDARs to TRPM4. This interaction is mediated by intracellular domains located in the near-membrane portions of the receptors. Structure-based computational drug screening using the interaction interface of TRPM4 in complex with NMDARs identified small molecules that spare NMDAR-induced calcium signaling but disrupt the NMDAR/TRPM4 complex. These interaction interface inhibitors strongly reduce NMDA-triggered toxicity and mitochondrial dysfunction, abolish cyclic adenosine monophosphate–responsive element–binding protein (CREB) shutoff, boost gene induction, and reduce neuronal loss in mouse models of stroke and retinal degeneration. Recombinant or small-molecule NMDAR/TRPM4 interface inhibitors may mitigate currently untreatable human neurodegenerative diseases.